Protocol 31820: Probe Assay - Slc2a7<em1(IMPC)J>
Version 1.0

Notes

Taqman qPCR protocols are run on a real time PCR instrument. Use an appropriate instrument specific Fluorophore/Quencher combination.

 

The genotyping protocol(s) presented here have been optimized for reagents and conditions used by The Jackson Laboratory (JAX). To genotype animals, JAX recommends researchers validate the assay independently upon receipt of animals into their facility. Reaction cycling temperature and times may require additional optimization based on the specific genotyping reagents used.

Expected Results

Mutant=  101 bp

Wild Type = 118 bp

>chr4:150149636+150149753 118bp GCTTCCACTTTCTTCCAGGAG TCCAGGTTCCCCTACACTGA

Sequence

Sequence (deleted region in lower case)

 

ACCAGATACAACCTCTTCACAACCCTACCTGTCCCTACACCTTCAAGGTCAAAAGCAGGAAAAATGTGTCCCTAGAAACAGGATAACACCTGCTTGGGTCACCTTCCCAGACAGCCCCCTCCTCCTCCTTCCAGGATTAGAGCCTGCTCCCAGCCAGACCTCCCTTTCCACctaaagaggaagtggaggcctggccaccctcaccccaccccaatcctagaagtgacaggagccaaccctacagagtctctgacctcgaaactctctggttcagcgtctgcagcccacactagtcctgactaccctaagtgctgcctttggctcagtcttccagtatggctacaacatcgccgtgatcaacacacctcacaaggtgggcacaaggtgggtctcagcgggtagaagtcagctgctgtctggcttctggaagcttccactttcttccaggagtcccctagatacctacccctaaggactaggggcatctagagccatTTTATCTAGCAATGTCATCTTTTCTAGGGGTATCAGTGTAGGGGAACCTGGATTCTCTGCCTGGGCACCCTGCCTTGCTCACTTGGGAAGGCCATTTGTGGGTGACAGACACATCAAGAATGAACTGTTAACTTTTAGAGATTGTGTTTGGTCAAAGAGAAGCAACAGAGATAAAGGCTGACTGGAGCAAGTTCA

324 bp deletion beginning at Chromosome 4 positive strand position 150,149,378 bp and ending after 150,149,701 bp (GRCm38/mm10).

JAX Protocol

Protocol Primers

Primer 5' Label Sequence 5' → 3' 3' Label Primer Type Reaction Note
37747 TCC TCC TTC CAG GAT TAG AGC Mutant Forward A
37748 TCC AGG TTC CCC TAC ACT GA Common A
37749 GCT TCC ACT TTC TTC CAG GAG Wild type Forward A
37750 Fluorophore-1 CCA GCC AGA CCT CCC TTT Quencher-1 MUT Probe
37751 Fluorophore-2 CTA CCC CTA AGG ACT AGG GGC Quencher-2 WT Probe

Reaction A

Component Final Concentration
Kapa Probe Fast QPCR 1.00 X
ddH2O
37747 0.40 uM
37748 0.40 uM
37749 0.40 uM
Wt Probe 0.15 uM
Mutant Probe 0.15 uM
DNA

Cycling

Step Temp °C Time Note
1 95.0 --
2 95.0 --
3 60.0 --
4 -- repeat steps 2-3 for 40 cycles
5 40.0 -- Forever
JAX uses a very high speed Taq (~1000 bp/sec), use cycling times recommended for your reagents.

Strains Using This Protocol

This is the only strain that uses this protocol.